7. Project Summary/Abstract Many industrial chemicals, environmental pollutants and other xenochemicals activate transcription factors belonging to the nuclear receptor superfamily, which leads to widespread genomic, epigenetic and transcriptional changes that disrupt key biological pathways and metabolic processes in liver and other tissues. The studies proposed focus on the liver nuclear receptor CAR (Constitutive Androstane Receptor; NR1I3), which is activated by structurally diverse xenochemicals and regulates transcription of hundreds of protein-coding genes important for processes such as xenobiotic metabolism, lipogenesis, glucose homeostasis, and inflammation, and has been implicated as a regulator of non-alcoholic fatty liver disease (NAFLD) development. We have discovered that xenobiotic agonists of CAR and other xenobiotic-responsive receptors induce or repress the transcription of several hundred nuclear-enriched long non-coding RNAs (lncRNAs) with epigenetic and gene regulatory potential, many of which have human orthologs. This proposal builds on these findings and on recent advances in liver cell zonation, single cell-based transcriptomic profiling, and gene co-expression network analysis to elucidate in an intact mouse liver model the effects of CAR-responsive lncRNAs on fatty liver disease induced by foreign chemical exposure. The studies proposed test the hypothesis that a subset of CAR-responsive lncRNAs control hepatic gene regulatory networks driving NAFLD and downstream pathologies, dysregulating processes such as lipid and carbohydrate metabolism, hepatic architecture and mitochondrial function in a liver cell type-specific and hepatic lobule zone-dependent manner. The work proposed uses TCPOBOP (1,4-bis[2-(3,5-dichloro-pyridyloxy)]benzene), a prototypic non- genotoxic chemical and CAR-specific agonist ligand, to address the seemingly paradoxical finding that persistent exposure to foreign chemical CAR activators induces NAFLD in mice fed normal chow diet, but suppresses NAFLD development in mice fed a high fat diet. These studies will elucidate the role of CAR, and the lncRNAs that it regulates, in fatty liver disease etiology and progression. Results obtained will give critical insight into the underlying mechanisms by which foreign chemicals dysregulate CAR-dependent metabolic pathways linked to NAFLD, which affects 25% of US adults and is a major cause of cirrhosis, hepatocellular cancer and liver failure. This work will refocus research efforts on xenochemical action to include mechanistic studies of single cell-based, spatially zonated gene regulatory networks and the non-coding transcriptome, and will serve as a paradigm for other foreign chemical-activated receptors that dysregulate gene expression in complex ways. Together, the proposed studies on CAR-responsive lncRNAs and their role in xenochemical- induced liver pathology may lead to new ways to prevent, diagnose or treat liver diseases induced by chemical exposure.